Interactions between protein surfaces guide virtually every cell-signaling pathway, and the discovery of molecules that perturb cell signaling represents a foremost goal of chemical biology. This goal is complicated by the reality that the domains that orchestrate signaling through protein-protein interactions are typically not enzymes, and lack deep, concave binding pockets. During the previous funding period we developed a strategy for designing molecules whose shapes complement the extended, shallow protein-interaction clefts on protein surfaces. This strategy is referred to as protein grafting and evolution, and the molecules are called miniature proteins. Our results indicate that miniature proteins bind protein (and DNA) surfaces avidly and very effectively discriminate their prescribed target from structurally unrelated proteins In this application, we expand and broaden the scope of protein grafting and evolution in two important ways, by (1) exploiting the aPP type II polyproline (PPII) helix (alone or in combination with the alpha helix) for recognition, and (2) by devising new strategies and technologies to analyze and select for homolog-specific ligands. We focus on the uniquely large class of protein domains that mediate cell signaling through interactions with proline-rich motifs First (Aims 1 and 2), we focus on discriminating between and among SH3 and EVH1 domains, two classes of cell signaling molecules of widespread fundamental and medicinal interest. To facilitate the discovery of optimally selective miniature protein ligands for these domains, we describe (Aim 3) a novel set of protein chips that permit (1) rapid analysis of miniature protein specificity during library evolution and (2) simultaneous selection for affinity and specificity on a sub-proteome-wide scale. We anticipate that the miniature proteins, we discover during this funding period will (1) reveal fundamental information about molecular recognition, design, and the complicated relationships between structure and activity, (2) provide new, unique tools in research, often facilitating experiments that would be difficult or impossible otherwise, and (3) identify therapeutic leads for the large number of non-enzymatic protein targets arising from the genome effort.